The type 2 secretion system (T2SS) is present in some Gram-negative eubacteria and used to secrete proteins across the outer membrane. Here we report that certain representative heteroloboseans, jakobids, malawimonads and hemimastigotes unexpectedly possess homologues of core T2SS components. We show that at least some of them are present in mitochondria, and their behaviour in biochemical assays is consistent with the presence of a mitochondrial T2SS-derived system (miT2SS). We additionally identified 23 protein families co-occurring with miT2SS in eukaryotes. Seven of these proteins could be directly linked to the core miT2SS by functional data and/or sequence features, whereas others may represent different parts of a broader functional pathway, possibly also involving the peroxisome. Its distribution in eukaryotes and phylogenetic evidence together indicate that the miT2SS-centred pathway is an ancestral eukaryotic trait. Our findings thus have direct implications for the functional properties of the early mitochondrion.

Biochemistry of Macromolecular Interactions Unit Department of Structural Biology and Chemistry Institut Pasteur CNRS UMR3528 Paris France

Center for Cellular Imaging and NanoAnalytics University of Basel Basel Switzerland

Department of Biochemistry and Molecular Biology and Centre for Comparative Genomics and Evolutionary Bioinformatics Dalhousie University Halifax NS Canada

Faculty of Science Department of Biology and Ecology University of Ostrava Ostrava Czech Republic

Faculty of Science Department of Genetics and Microbiology Charles University Prague 2 Czech Republic

Faculty of Science Department of Parasitology Charles University BIOCEV Vestec Czech Republic

Faculty of Science Department of Zoology Charles University Prague 2 Czech Republic

Faculty of Science Proteomic core facility Charles University BIOCEV Vestec Czech Republic

Institute of Molecular Genetics Czech Academy of Sciences Prague 4 Czech Republic

Institute of Parasitology Biology Centre Czech Academy of Sciences České Budějovice Czech Republic

Laboratory of Molecular and Evolutionary Parasitology RAPID group School of Biosciences University of Kent Canterbury UK

Robert Cedergren Centre for Bioinformatics and Genomics Département de Biochimie Université de Montréal Montreal QC Canada

School of Biosciences University of Birmingham Edgbaston UK

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